The long-term goal of this project is to characterize the relationship between dengue serotype 2 (DEN-2) viruses and their main vector, Aedes aegypti mosquitoes. Understanding this relationship will allow us to interrupt the transmission of this virus to susceptible human hosts, and thus decrease disease incidence. This is a competing continuation of grant AI50123, under which we derived the first comprehensive dengue phylogenetic trees, showed that distinct virus genotypes correlate with disease severity, identified probable viral virulence determinants, showed that these determinants are responsible for increased viral output by Southeast Asian viruses compared to American viruses, demonstrated differences in mosquito infection and dissemination rates, showed that selection for more virulent viruses occurs in both humans and mosquitoes, and developed a mouse model of dengue fever. In this continuation we propose to identify the molecular mechanisms by which SE Asian viruses get transmitted to humans at a higher rate than American genotype viruses, including identifying mosquito midgut viral receptors and whether mosquito saliva influences virus transmission in vivo. We will extend previous studies to actually reflect what occurs in nature. Dengue is a Category a Priority Pathogen in the NIAID Biodefense Research Program. This project would be integrated with our proposed R01 project concerning characterization of a mouse model of human dengue disease;the results of both projects could be integrated for a global strategy to control dengue, which is increasing in incidence, severity and geographic range. Since there are no treatments or vaccines for dengue, understanding how the vector selectively transmits more virulent viruses is very important. This project could help identify new ways to interrupt dengue virus transmission and epidemics. The specific aims are: 1. Characterize DEN-2 genotype-specific differences in infection, dissemination, and transmission rates in Aedes aegypti. Hypothesis: Structural differences between SE Asian and American genotype viruses determine dissemination and transmission rates by Ae. aegypti. 2. Determine differences in binding of DEN-2 viruses with Ae. aegypti midgut receptors and chemically characterize these receptor(s). Hypotheses: a. Differences in infection and dissemination between DEN-2 viruses are due to E glycoprotein differences, b. DEN-2 viruses use a specific mosquito midgut receptor to initiate an infection and this determines vectorial capacity. 3. Determine the effect mosquito saliva has on dengue virus infection and replication in a mouse model of dengue fever. Hypothesis: Aedes aegypti saliva facilitates and/or potentiates dengue virus infection of initial target (dendritic) cells.